The invention relates to a culture vessel which is adapted for culturing biological cells in hanging droplets, in particular a culture vessel in the inner space of which a culturing area is provided for receiving the hanging droplets. The invention also relates to a method for culturing biological cells in hanging droplets. Uses of the invention lie in the culturing of biological cells, in particular stem cells.
The cultivation (multiplication and/or differentiation) of biological cells in hanging droplets is a widely used culturing method. The biological cells can be cultured in the hanging droplets, wherein contact with solid substrate surfaces is avoided and a geometric arrangement of the cells similar to the arrangement of cells during their multiplication and differentiation in nature is realized. For the formation of the hanging droplets and their loading with the cells, the following techniques are conventionally known.
In a method which is typically performed manually, a flat bowl with a planar bottom, for example, the cover of a Petri dish is used. Initially, the cover is arranged on a support in such a way that an inner surface of the cover is exposed. Droplets of a cell suspension are placed on the surface with a pipette and subsequently, the cover is turned over by means of a pivoting movement such that the surface loaded with the droplets faces downwardly. The pivoting movement must take place so rapidly that the droplets do not run but remain in their places and, as a result, hang free on the downwardly facing inner surface of the cover after the pivoting. For the culturing, the cover is arranged, for protection against drying out, on the associated bottom of the Petri dish filled with an aqueous solution. This technique has the advantage of being simple to perform. However, disadvantages arise from the necessarily rapid and sudden pivot movement of the cover, which requires a high degree of skill from the user and permits automation to only a limited degree. Furthermore, undesired shear forces can arise in the droplets and these can have disadvantageous effects on sensitive cells, particularly stem cells.
Alternatively, hanging droplets can be generated on cell culture plates with holes which are surrounded at an underside of the plates by holding rings for holding the droplets by means of capillary forces. The formation of the droplets and their loading on one of the holding rings takes place in that the cell suspension is fed in through the associated hole in the plate and is hung as droplet on the holding ring (see also WO 2008/125347). The cell culture plate favors automation of the culturing method and the avoidance of the aforementioned shear forces. It is disadvantageous, however, that this technique demands special measures for preventing undesirable influences from the surroundings and mutual contamination of adjacent droplets.
It is an objective of the invention to provide an improved culture vessel which is configured for culturing biological cells in hanging droplets and is able to avoid disadvantages of conventional techniques. The culture vessel should enable, in particular, a gentle formation of the hanging droplets, offer protection of the hanging droplets and/or be automatable. It is a further objective of the invention to provide an improved method for culturing biological cells in hanging droplets, with which the disadvantages of conventional techniques are avoided. The method should, in particular, simplify the formation of the hanging droplets, wherein undesired shear forces can be minimized or excluded and/or enable the provision of pre-determined, reproducible culturing conditions.
These objectives are achieved by means of a culture vessel of the invention and a culturing method using the culture vessel, of the invention.